DESCRIPTION (adapted from the applicant's description): Familial Hypertrophic Cardiomyopathy (FHC) is an autosomal dominant disorder caused by mutations in any of several genes encoding the proteins of the cardiac contractile apparatus. This project will characterize the effects of FHC-causing mutations on the in vitro function of the various thin filament proteins so far implicated in this disorder: troponin T, troponin 1, and alpha-tropomyosin. By comparing normal and mutant proteins, the project will provide some of the insight required to understand the pathophysiology of cardiac disease in these patients. Also, the applicant will use the mutations to test the mechanism by which cardiac contraction is regulated by troponin and tropomyosin. A multi-faceted study of the mutant proteins is planned, with examination of several protein-protein affinities (including thin filament binding of troponin, of troponin-tropomyosin, and of myosin Sl; troponin binary subunit interactions; effects of calcium and of myosin on these various processes), calcium affinity, myosin MgATPase regulation, folding stability, in vitro motility, in vitro force, and structural effects on the regulatory conformational switching of the thin filament as determined by 3-D reconstructions of electron micrographs. A smaller number of mutations, identified in the cardiac actin gene, have been found causative in a subset of patients with another genetic disorder: dilated cardiomyopathy. Mutant forms of actin will be similarly examined for alterations in interactions with tropomyosin and troponin. (1) FHC mutations occurring in two regions of troponin T will be investigated. Troponin T mutants R92Q, R92W, Al 04V, and Fl 01 I occur in or near a region of troponin T that the applicant recently identified as forming a critical portion of the troponin tail. In a different region, the effects of FHC-linked COOH-terminal truncation of 28 residues will be studied. (2) Six troponin I mutations that occur in FHC will be similarly investigated, all located in the region of troponin I that interacts with the regulatory domain of troponin C. (3) Five FHC-linked tropomyosin mutants will be created and similarly studied, as will an actin mutation that causes inherited dilated cardiomyopathy and that is hypothesized to interact abnormally with tropomyosin.